EGFR is expressed in several solid malignancies, including NSCLC, HNSCC, malignant glioma and colorectal cancer, and abnormal or deregulated EGFR activity is known to contribute to numerous tumorigenic processes. Lung cancer remains the leading cause of cancer death in industrialized countries. Cancers that begin in the lungs are divided into two major types, non-small cell lung cancer and small cell lung cancer, depending on how the cells appear under a microscope. Non-small cell lung cancer (squamous cell carcinoma, adenocarcinoma, and large cell carcinoma) generally spreads to other organs more slowly than does small cell lung cancer. About 75 percent of lung cancer cases are categorized as non-small cell lung cancer (e.g., adenocarcinomas), and the other 25 percent are small cell lung cancer. For patients with advanced disease, chemotherapy provides a modest benefit in survival, but at the cost of significant toxicity, underscoring the need for therapeutic agents that are specifically targeted to the critical genetic lesions that direct tumor growth (Schiller J H et al., N Engl J Med, 346: 92-98, 2002).
Mutations that lead to EGFR overexpression (known as upregulation) or overactivity have been associated with a number of cancers, including lung cancer, anal cancers and glioblastoma multiforme. Mutations, amplifications or misregulations of EGFR or family members are implicated in about 30% of all epithelial cancers. Consequently, mutations of EGFR have been identified in several types of cancer, and has led to the development of anticancer therapeutics directed against EGFR, using two approaches: (1) targeted monoclonal antibodies (mABs) that prevent the binding of ligands to. EGFR, and (2) small molecule tyrosine kinase inhibitors (TKIs) that block the intracellular catalytic activity of the receptor. Skin toxicity characterized by rash or acne-like symptoms and diarrhea of different grades are the most common adverse events of EGFR targeted therapies (Expert Opin. Investig. Drugs (2009) 18(3), 293-300).
The human/mouse chimeric IgG1 mAb cetuximab down-regulates EGFR signaling and subsequently inhibits cell proliferation, induces apoptosis and reduces angiogenesis. Cetuximab in combination with chemotherapy has been approved by Health Authorities for the treatment of metastatic colorectal cancer and for the treatment of locally advanced and metastatic head and neck cancer. Cetuximab has also demonstrated little clinical activity as a single agent in patients with advanced NSCLC after prior EGFR TKI therapy (Neal J W, Heist R S, Fidias P, Temel J S, Huberman M, Marcoux J P, Muzikansky A, Lynch T J, Sequist L V; J Thorac Oncol. 2010 November; 5(11):1855-8: Cetuximab monotherapy in patients with advanced non-small cell lung cancer after prior epidermal growth factor receptor tyrosine kinase inhibitor therapy). Panitumumab (VECTIBIX®) is a human IgG2 mAB against EGFR and approved for treatment of metastatic colorectal cancer. Other monoclonals in clinical development are zalutumumab, nimotuzumab, matuzumab and necitumumab.
First generation small molecule HER TKIs include gefitinib (Iressa®) and erlotinib (Tarceva®), both binding reversibly to the EGFR. Gefitinib is indicated in all lines of treatment of advanced NSCLC harbouring EGFR mutations in the tumor and erlotinib is indicated as treatment of advanced NSCLC after prior chemotherapy, but in development in all lines of EGFR mutation positive NSCLC These new drugs directly target the EGFR. Patients have been divided into EGFR positive (EGFR+) and negative (EGFR−), based upon whether a tissue test shows a mutation. EGFR positive patients with tumors harboring EGFR mutations in exons 19 and 21 associated with drug sensitivity (i.e., G719X, exon 19 deletion, L858R, L861Q) have shown an response rate up to 60% which exceeds the response rate for conventional chemotherapy.
Second generation small molecule TKIs have been designed as irreversible EGFR inhibitors which bind irreversibly to EGFR, preferably to cysteine 773 of EGFR. Nonlimiting examples include compounds disclosed in U.S. Pat. No. 6,002,008, U.S. Pat. No. 7,019,012, U.S. Pat. No. 6,251,912, WO 02/50043, WO 2004/074263, WO 2005/037824, WO 2008150118 (specifically the compound of Example 36, 1-(4-(4-(3,4-dichloro-2-fluorophenylamino)-7-methoxyquinazolin-6-yloxy)piperidin-1-yl)prop-2-en-1-one, or salts thereof formed with acidic additives as disclosed in WO 2011155793), EKB-569 (pelitinib), HKI-272 (neratinib), HKI-357, CI-1033 (canertinib), WZ 3146, WZ 4002, WZ 8040 (structures of the three WZ compounds disclosed by Wenjun Zhou et al.: Novel mutant-selective EGFR kinase inhibitors against EGFR T790M, in Nature 2009, Vol. 462, 1070-1074), BIBW 2992 or PF-00299804. BIBW 2992 (afatinib) and PF-00299804 (dacomitinib) are most advanced second generation small molecule TKIs include, both in advanced clinical development for treatment of NSCLC.
More specifically, BIBW 2992, also referred to herein by it's INN afatinib, is known as the compound 4-[(3-chloro-4-fluorophenyl)amino]-6-{[4-(N,N-dimethylamino)-1-oxo-2-buten-1-yl]amino}-7-((S)-tetrahydrofuran-3-yloxy)-quinazoline,
preferably used as a maleate salt BIBW 2992:maleic acid 1:2:

BIBW 2992 is a potent irreversible and selective dual inhibitor of erbb1 receptor (EGFR) and erbB2 (Her2/neu) and erbB4 (Her4) receptor tyrosine kinases which can be administered orally. Furthermore, BIBW 2992 was designed to covalently bind to EGFR and HER2 thereby irreversibly inactivating the receptor molecule it has bound to. This compound, salts thereof such as the dimaleate salt, their preparation as well as pharmaceutical formulations comprising BIBW 2992 or a salt thereof, indications to be treated with BIBW 2992 and combinations including BIBW 2992 are disclosed in WO 02/50043, WO 2005/037824, WO 2007/054550, WO 2007/054551, WO 2008034776 and WO 2009147238.
PF-00299804 is an oral irreversible pan-HER TKI, more specifically an inhibitor of the HER1, 2, and 4 tyrosine kinases. In preclinical studies, PF-00299804 has been shown to inhibit the signaling in both wild-type and mutant EGFR, including forms of NSCLC that are resistant to currently available EGFR inhibitors, such as erlotinib and gefitinib. Preclinical findings suggest that PF-00299804 may be clinically effective against NSCLCs with EGFR or ERB-B2 mutations as well as those harboring the EGFR T790M mutation, which produces resistance to gefitinib and erlotinib (Expert Opin. Investig. Drugs (2010) 19(12): 1503-1514). PF-00299804 (dacomitinib) is the compound N-[4-(3-chloro-4-fluoro-phenylamino)-7-methoxy-quinazoline-6-yl]-3-piperidin-1-yl-acrylamide, disclosed in WO 2005107758 as Examples 2 and 3 with the following structure:

Characteristics of EKB-569 (pelitinib), HKI-272 (neratinib), HKI-357 and CI-1033 are published, e.g. Expert Opin. Investig. Drugs 2009, 18(3), 293-301 provides a review. Development of EKB-569 for treatment of NSCLC has been discontinued several years ago. Others report that HKI-272 can overcome T790M-mediated resistance only at suprapharmacologic concentrations (N. Godin-Heymann et al., The T790M “gatekeeper” mutation in EGFR mediates resistance to low concentrations of an irreversible EGFR inhibitor. Mol. Cancer Ther. 7, 2008:874-879). Remarkably, development of HKI-272 for treatment of NSCLC was discontinued after phase II trial showed that the compound had low activity in patients with prior benefit from TKIs and in TKI-naive patients, potentially because of insufficient bioavailability from diarrhea-imposed dose limitation (L. V. Sequist et al., J. Clin. Onc. 28 (18), 2010, 3076-3083). In contrast to encouraging preclinical results and high potency of HKI-272 these did not translate into clinical benefit, showing the low level of predictability in this field.
Despite initial response in NSCLC patients with EGFR mutations, acquired resistance develops after a median of approximately 12 months. The consensus definition of acquired resistance includes patients who had previous treatment with a single-agent EGFR-TKI (e.g., gefitinib or erlotinib); either or both of the following: a tumor that harbors an EGFR mutation known to be associated with drug sensitivity (i.e., G719X, exon 19 deletion, L858R, L861Q) or objective clinical benefit from treatment with an EGFR-TKI; systemic progression of disease applying RECIST criteria known in the art, while on continuous treatment with EGFR directed treatment for at least 24 weeks.
Response evaluation criteria in solid tumours (RECIST) are described by P. Therasse et al., J Natl Cancer Inst 2000, 92, 205-216; in J. Clin. Oncol. Vol 24, No. 20, 2006, pp 3245-3251; or by Eisenhauer E A, Therasse P, Bogaerts J, Schwartz L H, Sargent D, Ford R, et al., New response evaluation criteria in solid tumours: revised RECIST guideline (version 1.1). Eur J Cancer 2009; 45:228-247. Monitoring tumor progression may be determined by comparison of tumor status between time points after treatment has commenced or by comparison of tumor status between a time point after treatment has commenced to a time point prior to initiation of treatment. Tumor progression may be monitored during treatment visually, for example, by means of radiography, for example, X-ray, CT scan, or other monitoring methods known to the skilled artisan, including palpitation of the cancer or methods to monitor tumor biomarker levels.
In addition to the primary EGFR mutations (associated with erlotinib and gefitinib sensitivity), approximately half of the patients with acquired EGFR-TKI resistance have a second EGFR mutation (T790M) in the ATP-binding pocket of the tyrosine kinase that may alter receptor affinity in favor of ATP. These second mutations enable the cancer cells to continue signaling via mutant EGFR, suggesting that in a proportion of patients with acquired resistance to EGFR-TKIs, tumor growth and proliferation remains dependent on EGFR.
The presence of MET oncogene has been reported as a second sources of resistance (Jackman D, Pao W, Riely G J, Engelman J A, Kris M G, Janne P A et al., Clinical definition of acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors in non-small-cell lung cancer, J Clin Oncol 2010; 28:357-60).
As of 2010 there was no clinical consensus of an accepted approach to overcome or prevent resistance nor regulatory approval of a specific drug or drug combination in this setting.
There is a significant medical need in the art for a satisfactory treatment of cancer, and specifically epithelial cell cancers such as lung, ovarian, breast, brain, colon and prostate cancers, which incorporates the benefits of EGFR targeted therapy and overcoming the non-responsiveness exhibited by patients' cancers. Thus, the problem underlying the present invention is to establish an improved treatment of patients suffering from epithelial cell cancers, characterized by improved efficacy and improved or at least acceptable tolerability, including the following patient populations                (a) TKI naive cancer patients, wherein the improvement includes prevention or delay of resistance to TKI treatment,        (b) patients with tumors expressing the wild-type EGFR (described hereinbefore as EGFR),        (c) patients with tumors expressing mutated forms of the EGFR (described hereinbefore as EGFR+),        (d) patients previously treated with EGFR inhibitors, such as gefitinib or erlotinib afatinib, dacomitinib or others wherein the improvement includes to overcome primary or acquired resistance to EGFR inhibitors,        (e) patients with acquired resistance to treatment with TKIs such as gefitinib or erlotinib, afatinib, dacomitinib or others wherein the improvement includes to overcome resistance to TKI treatment,        (g) patient populations with primary or acquired resistance caused by T790M (T790M+), wherein the improvement includes to prevent/overcome resistance to TKI treatment, and        (h) patient populations with primary or acquired resistance not caused by T790M (T790M−), e.g. by other mechanisms such as MET oncogene or by unknown origin, wherein the improvement includes to prevent/overcome resistance to TKI treatment.        
One approach to improve treatment options of NSCLC patients with acquired resistance to gefitinib or erlotinib followed the concept of total receptor blockade by combining a TKI and an anti-EGFR mAB. This is summarized by S. Ramalingam et al., Journal of Thoracic Oncology Vol. 3, Number 3, March 2008, 258-265. The hypothesis was that by the combination it may be possible to achieve simultaneous vertical inhibition of EGFR and enhance abrogation of downstream activity. Residual EGFR activity after exposure to either class of inhibitor alone may allow cancer cells to remain viable, but simultaneous dual inhibition may cause apoptosis. Results in xenograft models support this hypothesis: a synergistic effect has been observed when cetuximab is administered in combination with erlotinib or gefitinib compared with treatment with either agent alone. Cetuximab has been shown to down-regulate EGFR on the cellular surface, potentially enhancing the sensitivity to TKIs. The conclusion was that the combination of cetuximab plus erlotinib seems synergistic in terms of apoptotic activity in vitro, and results in additive tumor growth inhibition in vivo.
Nevertheless, the question is whether these primary results translate in clinical benefit. S. Ramalingam et al. report the results of a phase I study carried out to determine the optimal doses of cetuximab and gefitinib when administered as a combination for patients with advanced/metastatic non-small cell lung cancer (NSCLC) previously treated with platinum-based chemotherapy. Patients with advanced/metastatic NSCLC treated with prior platinum-based chemotherapy received escalating doses of weekly cetuximab (100, 200, and 250 mg/m2, iv) and fixed doses of gefitinib (250 mg/d, PO) until disease progression or unacceptable toxicity. The results reported show that the combination of cetuximab and gefitinib can be safely administered but has only modest activity in advanced/metastatic NSCLC.
Y. Y. Janjigian et al., Clin Cancer Res 2011, 17: 2521-2527, report about a phase I/II trial of cetuximab and erlotinib enrolling 19 patients with lung adenocarcinoma and acquired resistance to erlotinib. Patients with lung adenocarcinoma and clinically defined acquired resistance to erlotinib were treated with erlotinib 100 mg daily, along with cetuximab every 2 weeks in three escalating dose cohorts (250 mg/m2, 375 mg/m2, and 500 mg/m2). The recommended phase II dose was then evaluated in a two-stage trial, with a primary, end point of objective response rate. The recommended phase II dose identified was cetuximab 500 mg/m2 every 2 weeks and erlotinib 100 mg daily. At this dose and schedule, no radiographic responses were seen. In fact, combined EGFR inhibition, with cetuximab 500 mg/m2 every 2 weeks and erlotinib 100 mg daily, had no significant activity in patients with acquired resistance to erlotinib. During the phase II portion of the trial serious tolerability issues occurred. Common grade 2, 3 and 4 toxicities were rash (13 patients, 68%), fatigue (12 patients, 63%) and hypomagnesemia (14 patients, 74%). 31% (6 of 19 patients) discontinued treatment due to intolerable rash.
Both, S. Ramalingam et al. and Y. Y. Janjigian et al., report clinical results obtained with the combination of cetuximab with a reversible (first generation) TKI. In contrast, L. Regales et al., J. Clin. Invest. 119 (10), 2009: 3000-3010, report results obtained with the irreversible (second generation) TKI BIBW 2992 in transgenic mouse lung tumor models that develop lung adenocarcinomas driven by EGFRL858R (sensitive to erlotinib), EGFRT790M (resistant to erlotinib), or EGFRL858R+T790M (resistant to erlotinib) with a focus to evaluate strategies to overcome the most common EGFR TKI resistance mutation, T790M. Other agents mentioned in the investigation were HKI-272 (neratinib) and PF-00299804, but without reporting results. The rationale behind was that preclinical studies published by others (E. L. Kwak, et al., Irreversible inhibitors of the EGF receptor may circumvent acquired resistance to gefitinib. Proc. Natl. Acad. Sci. U.S.A. 2005, 102:7665-7670; T. A. Carter et al., Inhibition of drug-resistant mutants of ABL, KIT and EGF receptor kinases. Proc. Natl. Acad. Sci. U.S.A. 2005, 102:11011-11016) suggested that second-generation irreversible EGFR inhibitors may be able to overcome T790M-mediated resistance, at least in vitro. Mice bearing tumors harboring EGFR mutations were treated with a variety of anticancer agents, including the irreversible EGFR TKI BIBW 2992 and the EGFR-specific antibody cetuximab. It was found that only the combination of both agents together induced dramatic shrinkage of erlotinib-resistant tumors harboring the T790M mutation.
An open label phase I clinical trial of continuous once daily oral treatment using BIBW 2992 in combination with cetuximab with the primary objective to determine the maximum tolerated dose (MTD) and recommended phase II doses in patients with NSCLC and acquired resistance to erlotinib or gefitinib was disclosed in ClinicalTrials.gov at the priority filing date of the subject patent application, identifier NCT01090011, including the history of changes available via a link to ClinicalTrials.gov archive site. The following qualitative information about the administration regimen was publicly available:                patients to receive medium BIBW 2992 once daily plus biweekly cetuximab infusion at low, median and high dose level        BIBW 2992 medium dose plus three dose levels (low, medium and high) of cetuximab.        
Results or absolute dosages were not disclosed.